1. Field of the Invention
The present invention relates generally to novel antitumor compositions and their use in treating tumor cells. More particularly, the invention is directed to a soluble antitumor factor released by precisely activated cells of monocyte/macrophage lineage and further to methods for its purification and use.
2. Description of the Related Arts
Recently, great interest has been shown by medical scientists in the identification and isolation of a variety of biologically-derived factors which have shown promise in the treatment of various disease states. Moreover, a number of such biological factors have been identified that demonstrate cytotoxic anticellular activities which are selective for neoplastic cells, thus holding the promise that such factors may provide additional weapons in the anti-cancer armamentarium.
Macrophages have been a particularly important focus of attention as an appropriate arm of host response that might be manipulated for the immunotherapy of neoplastic disease. This attention is warranted because of the ubiquitous distribution of these cells in tissues, and because of their demonstrated ability in vitro to mediate a lytic mechanism preferentially manifested on tumor cells (See, e.g., Fidler (1978), Isr. J. Med. Sci., 14:177). More recently, the activation of macrophages by targeted agents in vivo has led to the eradication of metastatic foci in animal model systems (See, e.g., Fidler et al. (1982), Cancer Res., 42:496; Lopez-Berestein et al. (1984), Clin. Exp. Metastasis, 2:127).
Characterization of the tumoricidal mechanism has been conducted by a number of laboratories, and a striking scope of pathways has been attributed to the effector cell. Although there is widespread agreement that under most circumstances the lytic mechanism is facilitated by contact between the macrophage and the target cell, in many systems this has been shown not to be obligatory.
In the human system, reports from several laboratories have described macromolecular cytotoxins derived from human monocytes and macrophages. For example, Reed and Lucas ((1975), J. Immunol., 115:795) have described a 45 kilodalton toxin that arose apparently spontaneously after adherence of peripheral blood mononuclear cells. Matthews, in a 1981 report, demonstrated that adherent peripheral blood monocytes could release a cytotoxin after 20 hours of incubation with varying levels of endotoxin triggering of the monocytes. ((1981), Immunol., 44:135). This toxin exhibited a molecular weight of 34 kilodaltons by molecular sieving chromatography.
In contrast to the results of Reed and Lucas and Matthews, Hammerstrom found that peripheral blood monocytes from BCG-vaccinated donors required up to 3-5 days of total in vitro incubation and 24 hrs. incubation with crude human lymphokine preparations to release cytostatic factors. ((1982), Scand. J. Immunol., 15:331). Previous studies by Hammerstrom had shown that this protocol produced competent effector cells for direct tumor cell cytotoxicity. In further studies, two cytostatic factors were characterized and partially purified by Nissen-Meyer and Hammerstrom ((1982) Infect. Immunity, 38:67), and found to exhibit molecular weights of about 40 and 55 kd.
In 1983, Cameron reported that cytotoxic activity could be obtained spontaneously from peripheral blood monocytes cultured in vitro for five days ((1983), J. Reticuloendo. Soc., 34:45). This factor(s) was shown to enhance host survival in animal tumor systems. However, little molecular characterization was conducted. In 1984 Sone et al. ((1984), Cancer Res., 44:640), found that human alveolar macrophages could be triggered with LPS (lipopolysaccharide) or MDP (muramyl dipeptide) to release a cytolytic factor. The ability of alveolar macrophages to release the factor was found to persist for up to 48 hours. However, as with the studies of Cameron, no biochemical characterization of the factor was reported.
A number of human cell lines have been exploited as models of monocytes/macrophages. Some of these have been found to release cytotoxic macromolecules following appropriate triggering. For example, Gifford et al. ((1984), J. Natl. Cancer Inst., 73:69) demonstrated that KG-1, HL-60, ML-2, and ML-3 lines could be induced with PMA (phorbol myristate acetate) to release cytotoxins; the principal species from the ML-2 line was found to be 40 kilodaltons. Armstrong et al. ((1985), J. Natl. Cancer Inst., 74:1) used the THP-1 cell line which, following exposure to relatively high levels of PMA, elaborated primarily a 120 kilodalton cytotoxic species believed to be a neutral protease. Pennica et al. ((1985), Nature, 312:724) reported the cloning of a TNF alpha cDNA which apparently coded for about a 17 kilodalton protein.
Therefore, in general, the prior art has demonstrated the existence of numerous biological response modifiers, certain of which have shown promise as potentially useful clinical agents. Moreover, numerous such factors of monocyte/macrophage origin have been described and characterized to varying extents and are presently being investigated to determine their efficacy in treating tumors. The present invention is directed to a novel antitumor factor isolated from cells of macrophage/monocyte lineage which has been characterized both in terms of its biological and biochemical attributes. It is believed by the present inventors that this novel factor will provide an additional tool for the medical oncologist in the treatment of various tumors.